Molecular Switch Regulates Fat, Cholesterol Metabolism

Doctors typically treat hypertension, type 2 diabetes, obesity, non-alcoholic fatty liver, and atherosclerosis individually, though all may involve dysfunctional cholesterol or fat homeostasis. Many of these conditions have recently been categorized as components of metabolic syndrome, a constellation of related diseases.

In the Aug. 10 issue of Nature, researchers at HMS and the Massachusetts General Hospital Cancer Center describe the interaction of several key regulators involved in the body’s control of lipid levels. “The identification of this protein interaction and the nature of the molecular interface may one day allow us to pursue a more comprehensive approach to the treatment of metabolic syndrome,” said Anders Näär, principal investigator and HMS assistant professor of cell biology at the MGH Cancer Center.

Image courtesy of Anders Näär

Fat stores. Anders Näär and colleagues identified an interaction between transcription regulators SREBP and ARC105 that controls production of enzymes that allow C. elegans to store fat by converting stearic to oleic acid. Fat storage (indicated by dark intestines in a healthy worm, left) is lost with inhibition of worm SREBP or ARC105 (middle, clear intestines) and restored in worms fed dietary oleic acid (right).

Between meals, cholesterol and fat production should be turned off, but excess food intake and lack of exercise appear to disturb the normal checks and balances that control the sterol regulatory element binding protein (SREBP) transcription factor family, resulting in overproduction of lipids. Näär and colleagues studied how these SREBPs turn on genes involved in lipid synthesis. The researchers identified ARC105, a subunit of the large ARC/Mediator coactivator, as a critical effector of this SREBP gene regulation and revealed the precise interaction between these two molecules.

One domain of the SREBP protein grips DNA while a flexible tail extending from its other side fits into a groove on the ARC105 protein, allowing gene transcription to occur. The research group of Gerhard Wagner, the Elkan Blout professor of biological chemistry and molecular pharmacology at HMS, found this binding site by exhaustively testing fragments of ARC105 using NMR spectroscopy until they found a sequence, the KIX domain, that interacts with SREBP.

Malfunction of either SREBP or ARC105 has steep consequences. When Näär and the worm genetics group of Anne Hart, HMS associate professor of pathology at the MGH Cancer Center, knocked down either of the SREBP or ARC105 homologs in C. elegans using RNA interference (RNAi), the worms lost motility, fertility, and the ability to store fat. The researchers traced these phenotypic defects to a depletion of fat-6/fat-7, which transform stearic to oleic acid. The scarcity of these enzymes resulted in a buildup of stearic acid and depletion of oleic acid.

When researchers directly blocked function of the fat-6/fat-7 enzymes, similar phenotypes occurred—poor fat storage, infertility, and disabled motility. Feeding oleic acid to the worms resulted in partial recovery from many of these defects, suggesting the fat-6/fat-7 malfunction was the immediate cause of the worms’ disabilities. At a higher level of regulation, phenotypic changes depend on SREBP and ARC105, which like assembly line overseers, switch production of these enzymes on or off from their control room, determining how much oleic acid gets churned out below them on the factory floor.

This new understanding of SREBP and ARC105 suggests that their interaction might be a target for therapeutic intervention in the treatment of diseases associated with metabolic syndrome. The researchers have begun high-throughput screening at the ICCB-Longwood Screening Facility to identify small molecule inhibitors of the KIX binding site. Therapeutic applicability, though, remains a distant goal, cautions Näär. “There are numerous hurdles that would need to be overcome before finding specific and effective treatments based on these findings.”

—Kathleen Fink with reporting by John Lacey

Corneal Transparency Made Clearer

The cornea of the eye remains a pristine oasis of clarity in the snarled mesh of blood vessels that riddle the rest of the body. Vision depends on the cornea enforcing this rigorous ban on blood vessel growth, but how it does this remains poorly understood. “This phenomenon—avascularity of the cornea—has really been a feature of the cornea that has been puzzling for millennia,” said Reza Dana, of the Schepens Eye Research Institute, Massachusetts Eye and Ear Infirmary, and an HMS associate professor of ophthalmology. He and colleagues have uncovered a key mechanism keeping the cornea clear. In the July 25 Proceedings of the National Academy of Sciences, they build a case for the crucial involvement of vascular endothelial growth factor receptor 3 (VEGFR-3) in maintaining corneal transparency.

VEGFR-3 appears to act as a sink for pro-angiogenesis signals, luring mediators VEGF-C and VEGF-D to bind this receptor rather than activating other pathways (by binding VEGFR-2) that promote blood vessel growth. Dana and colleagues determined that expression of VEGFR-3 occurs on cells of the epithelial layer, those making up the outer surface of the cornea, by using immunohistochemical staining and quantitative PCR on mouse corneal cells. This expression of VEGFR-3 in a location “not thought to be a natural home to this receptor, in itself was a novel observation,” said Dana.

They moved to in vivo mouse experiments using application of mild or more robust inflammatory stimuli to induce corneal inflammation. Both intact corneal epithelium and isolated corneal epithelium dampened recruitment of inflammatory cells and blocked the growth of new blood vessels. In corneas with the epithelium removed, however, significant vessel growth occurred. In addition, the researchers showed that these inflammatory insults to the cornea caused upregulation of pro-angiogenic factors VEGF-C and -D.

Two final experiments tested whether VEGFR-3 could be implicated as the specific factor making the corneal epithelium so resistant to blood vessel growth. The researchers removed corneal epithelium, and then introduced VEGFR-3 alone, which reinstated the cornea’s capability to block angiogenesis. Conversely, in mice with intact corneal epithelium, blocking VEGFR-3 resulted in inhibition of the epithelium’s anti-angiogenic capacity.

The results paint a picture of VEGFR-3 functioning as a sink in the corneal epithelium, sequestering pro-angiogenic mediators VEGF-C and -D so they cannot bind other receptors promoting angiogenesis. Redundant systems and additional mediators may also play a role in maintaining the cornea’s hallmark transparency, but it is clear that “this function of the epithelium is critically dependent on expression of VEGFR-3,” said Dana.

Halting angiogenesis by overexpressing VEGFR-3 may aid conditions of the eye in which blood vessel growth is pathogenic, such as corneal vascularization, the second leading cause of blindness in the world, and age-related macular degeneration, which causes blindness due to uncontrolled blood vessel growth at the back of the eye. Applicability of a VEGFR-3 sink may also extend to nonocular uses such as tumor suppression. “We’ve had a lot of positive feedback from oncologists,” said Dana.

—Kathleen Fink

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